The purpose of this project is to develop a protocol for rescuing victims from otherwise lethal (900cGy) total body irradiation (TBI). The protocol development will done using the preclinical dog model with umbilical cord blood as a source of stem cells. Cord blood transplantation results in less severe graft-versus-host disease (GVHD) compared to other sources of hematopoietic cells despite greater major histocompatibility complex (MHC)-disparity between the unrelated donor and recipient. A partially-MHC-matched cord blood unit can be rapidly identified for a large number of patients including members of minority ethnic/racial groups. However, for successful transplantation of cord blood into adults, a major problem is the low number of hematopoietic cells contained in cord blood units. This results in a very prolonged time to recovery of granulocytes and platelet counts and a high risk of graft rejection and mortality. Therefore, this project will focus on practical interventions to prevent rejection and enhance rapid engraftment of low cell number, partially-MHC-matched cord blood using the well-established dog model of hematopoietic cell transplantation. To facilitate engraftment of a low cell dose umbilical cord/neonatal blood (UCNB) graft, we will study the co-infusion of three distinct hematopoeitic cell sources that can be stockpiled and used "off-the-shelf". First, we will supplement the UCNB graft with 1, 5 or 10 units of cryopreserved, MHC-mismatched UCNB. Second, we will supplement low cell dose UCNB grafts with committed myeloid progenitor cells (from Project 4). Third, we will ex vivo expand UCNB derived CD34+ progenitor cells using the ligand for Notch-1 receptor. We will compare transplantation of expanded CD34+ UCNB cells alone and also in combination with a low cell dose UCNB graft. In all cases postgrafting cyclosporine and mycophenolate mofetil will be given to prevent GVHD. If GVHD persists, new strategies will be incorporated as developed in Project 6. Once conditions are achieved for reliable engraftment, the "window of opportunity" during which UCNB hematopoietic cells can rescue myeloablated recipient dogs will be determined. Completion of these pre-clinical studies in the dog model will be a major contribution to improving the outcome of patients who require ultra-urgent rescue of hematopoiesis after marrow-lethal TBI.